Shri Adhithya.

XII-A
 BONAFIDE CERTIFICATE

This is to certify that the Chemistry investigatory project on the topic

“_____________________________________________” has been
successfully
completed by ________________________ of class XII –____ Roll No:

___________ at FIITJEE GLOBAL SCHOOL Vengambakkam for the

partial fulfillment of this project as a part of All India Senior Secondary
Certificate Examination CBSE, New Delhi for the academic year 2024-2025.
DATE: _________________

Signature of Principal Signature of Guide

Submitted for SSCE 2024-2025 Chemistry Practical Examination held on

__________

INTERNAL EXAMINER EXTERNAL EXAMINER

 ACKNOWLEDGEMENT

I hereby express my heartfelt thanks to The Principal Ms. S MOHANA PRIYA who
has given me the opportunity to do the project in the school’s Chemistry laboratory and
for her constant encouragement.

I extend my heartfelt gratitude to Ms. Ashwathi, PGT Chemistry, for his invaluable
guidance and unwavering support. His encouragement, inspiration, and insightful
advice have been instrumental in the successful completion of this project.

I am also thankful to all my teachers and non-teaching staff for their help during my
course of study.

With heartfelt gratitude, I extend my deepest thanks to my parents for their unwavering
support and encouragement.
4 | Page
 STATEMENT OF PROBLEM

Casein is an essential protein. It provides the body with amino acids necessary
to build muscles. As we require a certain amount of muscle to facilitate the
normal functioning of the body, it is important to identify which sources of
milk help with muscle growth. A question that might arise to your mind might
be: -
Does cow milk, which is regularly consumed, help with bulking up better than
milk from other sources?
We will attempt to answer this question through an experiment to determine
the amount of casein in various kinds of milk.

OBJECTIVES
To know the role of casein in our body

To examine the relationship between casein and muscle growth To

determine the amount of casein in various types of milk

5 | Page
 INTRODUCTION

WHAT IS CASEIN?

Casein is a family of related phosphoproteins . found in milk that gives milk its
white colour. In pure form, it is an amorphous white solid, tasteless and
odourless without taste or odour, while its commercial type is yellowish with a
pleasing odour. Take cottage cheese, for example — the liquid settled on top
contains whey that has separated from the firm casein-containing curds below.
Newly opened or unmixed yogurt is another example of when you can often
see the whey-containing liquid separated from the solid yogurt. Cow's milk
consists of around 80% casein protein and between 20% and 60% of the
proteins in human milk. In addition to milk, casein protein is found in yogurt,
cheese, and infant formulas, as well as in a variety of dietary supplements. Do
not confuse casein protein with casein peptides. Casein peptides are made by
breaking casein protein down into smaller pieces. Structure and in nature,
casein exists as a molecule that's suspended in a surrounding liquid. This
structure is called a micelle. You may picture a micelle as an intact little bubble,
mixed into a solution. Casein is the principal milk protein. It exists in four
subtypes:

• αs1-casein
•
αs2-casein
•
• β-casein
• κ-casein
And α-Casein (a mix of αs1- and αs2-casein) is the main fraction of milk
protein.

6 | Page
Casein is the most important protein component in milk, both quantitatively
and nutritionally, accounting for about 80% of milk’s total nitrogen. It was used
in industries producing paper, textiles, paint, leather, fibre, and other
materials. Edible casein and caseinates are also long-established dairy
byproducts with uses in many foods. Casein is a very rich source of essential
amino acids, with the only possible exception of cysteine. It is a
phosphorylated and glycosylated complex synthesized by mammary glands. It
is constituted of three different polypeptide chains (αs1, αs2, and ß) held
together by noncovalent interactions. Casein fractions are organized in micellar
aggregates that also contain bivalent cations (calcium and smaller amounts of
magnesium), ranging 20–300 nm in diameter. This structure allows highly
stable dispersion of hydrophobic fractions in a colloidal state by the action of
hydrophilic bonds.

The amount of casein in whole milk varies according to the animal breed and
stage of lactation. It is generally in the range 24–29 g l− 1. Casein contains 0.7–
0.9% phosphorus, covalently bound to the protein by a serine ester linkage.
Consequently, casein is known as a phosphoprotein. All the amino acids that

7 | Page
are essential for humans are present in casein in high proportions, with the
possible exception of cysteine.

Thus, casein is considered to be a highly nutritious protein. It exists in milk in

complex groups of molecules called as micelles. The micelles consist of casein
molecules, calcium, inorganic phosphate, and citrate ions, and they have a
typical molecular weight of several hundred million daltons.

In terms of physical chemistry, the casein micelles exist in milk as a very stable
colloidal dispersion. As a protein, casein is made up of hundreds of individual
amino acids, each of which may have a positive or a negative charge,
depending on the pH of the milk system. At some pH value, all the positive
charges and all the negative charges on the casein remain in balance (i.e., the
net charge on the protein is zero); this pH value is known as the isoelectric
point (IEP), which is 4.6 for casein. The IEP is the pH at which the protein is
least soluble. Milk has a pH value of about 6.6, at which the casein micelles
have a net negative charge and are quite stable. Casein consists of several
individual casein components (αs1-, αs2-, β-, and κ-casein), each having slightly
different properties.

Casein is precipitated from skim milk by acidifying it to produce acid casein, or

the milk is treated with rennet to produce rennet casein. The precipitated
casein curd is separated from the whey, washed, and dried. The water-soluble
derivatives of acid caseins, produced by reaction with alkalis, are called
caseinates. Edible casein is a long-established dairy by-product used as an
ingredient in many food products, including dairy products. The general
development of food technologies and their applications has increased the
production of and demand for casein. Its manufacture differs from that of

8 | Page
nonedible casein (also called industrial casein) in that casein for food is
produced under sanitary conditions.

Further, during its manufacture, food-grade chemicals are used and sufficiently
heat-treated to make the casein safe for human consumption. The intensive
investigation into manufacturing technologies over the years and the
introduction of efficient plant designs have immensely improved the
technology for edible casein production.

SOURCES OF CASEIN:

1. Cow's milk, goat’s milk and buffalo milk

2.Curds and Cottage Cheese

Cottage cheese is made by clotting skim milk with rennin or lactic acid. Cottage
cheese is more protein-dense than milk, with 1 cup containing almost 27 grams
of protein, giving it a higher casein content than you'd get in milk. The protein
content of creamed cottage cheese will be lower, however, as the fat content
is ratcheted up to 4 percent, thus reducing the protein content a little.

ROLES OF CASEIN:

1. Growth and Development

Casein is a complete protein that contains all of the essential amino acids
required by the body. Each subunit of casein possesses a different sequence of
amino acids, lending to its diverse amino acid makeup. Because of its rich
amino acid profile, scientists have assumed that its role is to provide a source
of amino acids for the growth and development of infants, children and young
animals. It is an especially good source of lysine, an amino acid that is often

9 | Page
low in grain-based diets. However, another one of casein's important purposes
may be to prevent calcification of the mammary gland.

2. Health and Digestion

Casein is popular among athletes and bodybuilders due to its reputation as a

muscle builder. Casein protein is taken by mouth to improve athletic
performance, diabetes, liver disease due to alcohol consumption, and many
other conditions. In fact, casein supplementation was shown to increase serum
amino acid levels and muscle building after strength training in a study
published in 2004 in the journal, "Medicine and Science in Sports and
Exercise." When consumed, the casein micelle is disrupted by acid in the
stomach, which forms a clot and allows the slow breakdown of the protein
and, therefore, a sustained supply of amino acids over several hours, allowing
the body to retain and use those amino acids more effectively.

3. Food Technology

Cheese-makers also take advantage of the clotting features of casein. Enzymes

disrupt the micelle and cause the casein to clot -- and the clot is separated out
to form the curd. The curd is then further processed into cheese. Additionally,
the properties of casein make it possible to remove the fat from milk. Milk can
be centrifuged, or spun at extremely high speeds, which separates the milk
into a cream layer, a watery layer called a supernatant, and a solid layer called
a pellet. The supernatant is known as skim milk. The casein micelle is the
centre piece to the many functions of casein.

4. Industrial uses

10 | Page
  Paper coatings: Book and art papers are coated with pigmented casein

to provide a suitable surface for half-tone illustrations.

 Glues: Wood joints made with casein glue withstand dampness for some

time

CASEIN AND MUSCLE STRENGTH:

Casein helps muscle breakdown by its protein synthesis. For muscles to grow
they need to undergo protein synthesis. This process is the basis for muscle cell
growth and is essential for repair and maintenance of cells as well. This process
is increased when the muscle experiences a stimulus, such as the load they
would take on in a resistance training program. Consumption of adequate
amounts of protein is a necessary requirement for muscle protein synthesis,
the maintenance of muscle mass and keeping the muscles working properly.
Protein synthesis of the muscles must always be greater than muscle protein
breakdown. Numerous studies that have measured this balance between
protein breakdown and protein synthesis have found the balance to be
negative until protein was taken in through the diet, clearly stating the
important role protein plays in muscle protein synthesis. Protein synthesis is an
ongoing process that is constantly occurring in the body; however, it increases
drastically when resistance training is paired with a diet of protein. This
repeating turnover of protein is how damaged cells from exercise are turned
into new healthy cells. Unfortunately, this system isn’t completely full proof in
the recycling of every amino acid in their entirety. During this process many
amino acids are lost from the skeletal muscle and typically end up going one of
two routes. They are either oxidized or go through gluconeogenesis, which is
the process of converting the amino acids into glucose. These losses in amino

11 | Page
acids is where extra protein in the diet may be needed to make up for what is
lost during this process. This covers a little more deeply the ongoing need for
protein.
Casein may be beneficial to use before going to bed, allowing the amino acids
to help recover muscles while one sleeps.

BENEFITS OF CONSUMING CASEIN RICH MILK:

Casein “lasts” longer in your system

Not all proteins are created equal. Whether it’s soy, egg, whey, animal, or
casein protein, they all have their own unique advantages. Arguably, casein’s
greatest strength is timing. Casein has the ability to provide your bloodstream
with a slow and steady flow of amino acids that could last for hours. Muscles
may not be built overnight, but drinking a glass of casein-rich milk is the ideal
protein to consume right before bed, as it’ll be more helpful throughout the
night than any other protein option.

Greater Fat Loss

Since casein protein is a form of protein that is higher in calcium content that
also proves to be a benefit in terms of total fat loss. Many individuals are quick

12 | Page
to turn away from dairy products while attempting to lose body fat because
they feel it will slow them down.

This is completely opposite from the truth. One study that was conducted by
the International Journal of Obesity observed that those who combined a high
calcium intake over a twenty-four-hour period with a normal protein intake
showed increased faecal fat and energy excretion for that day of
approximately 350 kJ greater than those who either took in a low calcium,
normal protein intake or those who consumed a high protein, high calcium
intake.

This study makes mention of the importance of total protein intake with this
factor, stating that it appears as though moderate levels of protein create this
effect with the calcium greater than very high protein intakes do.

It appears from this research that it's best to find a good balance between
calcium and protein intake, rather than just increasing protein up higher to
promote fat loss.

Casein protein will help you accomplish both goals of having a higher calcium
intake as well as boosting your protein intake, allowing you to maximize your
fat loss benefits.

Helps Promote Colon Health

Another very important benefit of casein protein is that it helps promote colon
health. In a study performed out of Australia, researchers investigated the
health benefits of various proteins and found that dairy proteins promote
colon health better than meat and soy. The colon is a major organ essential for

13 | Page
waste removal in the body. If the colon is not working properly, the body can't
absorb essential nutrients or rid itself of waste. Lifestyle choices and eating
habits contribute to your colon's overall health.

It promotes dental health

Research says, one of the benefits of consuming casein is that it prevents

dental problems, as they have properties to reduce or prevent the effects of
enamel erosion.

It is a superior quality of protein

Casein is one of the best sources of high-quality protein available. It helps you
meet your minimum protein requirements by giving maximum quality of
protein, leading to lesser needs to maintain lean muscle mass.

COMPARING CASEIN AND WHEY PROTEIN:

Casein protein is good for various reasons:

It can provide your bloodstream with a steady flow of amino acids that will last
for hours.

When combined with whey, it is the best supplementation for achieving lean
muscle mass.

It boosts your metabolic rate and increases your level of satiety by 33%, which
is the easiest way to grow muscle and lose fat;

It has anti-catabolic effects (prevents muscles from breaking down) for more
muscle strength.

14 | Page
It is good for your teeth, as it reduces the chances of eroding your enamel. The

negative side of casein protein consumption is that it’s allergenic. People

that are allergic to milk may experience an allergic reaction. It can also cause
you to bloat, due to the sodium it contains.

On the other hand, Whey is known as a faster protein, and the main result you
get from it is faster protein synthesis, meaning the metabolic process of
protein is somewhere around 20 minutes (compared to casein, which is 3-4
hours). In about an hour, whey will be running through your bloodstream.

But whey is generally not preferred as it affects an individual’s immune

function and also there are no results proving that whey is effective in protein
degradation.

HYPOTHESIS
The main aim of this project is to determine and find out the amount of casein
in different sources of milk. The hypothesis we propose is that the amount of
casein decreases in the order:

Cow milk > Buffalo milk > Goat milk

We infer this because cow milk is very widely and commonly used than buffalo
and goat milk, because of its high nutritional value mainly being protein and
also because it is more economical.

Our research is based on an experimental investigation. The experiment was

carried out successfully.

15 | Page
 PROBLEM QUESTION
To study the amount of casein present in different samples of milk and thereby

conclude which type of milk is the most efficient.

EXPERIMENT

Aim:
To determine the amount of casein present in different samples of milk.

Apparatus required: Beakers, funnel, glass rod, weighing scale, test tubes,
filtration flask, Bunsen burner,100ml cow milk, 100ml buffalo milk,100ml goat
milk, 10% acetic acid solution, filter paper, 40ml ethanol, thermometer,
dropper and watch glass.

Theory:

16 | Page
Milk is a multinutrient fluid and it is the primary source of nutrition for human.
It consists of 80% of proteins. The protein in the milk is classified into casein
and whey protein. Milk protein consists of 80% of casein and 20% whey
protein. The function of casein is to provide energy to the human body. The
name of casein is related to the family of phosphoproteins. These proteins are
commonly found in mammalian milk. This study deals with the precipitation of
casein from the various milk samples such as cow milk, goat milk, and buffalo
milk. The technique of precipitation of casein is used to predict the protein
content in the milk samples.

Casein is the main protein constituent of milk. It constitutes about 80% of the
total protein in cow’s milk and about 3% of its weight. Its groups of protein
precipitated when the milk is slightly acidified. It dissolves slightly in water,
extensively in alkalis or strong acids. Casein is a complete protein meaning that
it contains all of the essential amino acids, which the body cannot manufacture
on its own. When dried, it is a white, amorphous powder without taste and
odour. It is a mixed phosphoprotein and occurs in milk as calcium salt (calcium
caseinate) in the form of micelle. The micelle has a negative charge. When an
acid is added to the milk, the negative charges are neutralized.

Calcium caseinate + acetic acid → casein (s) + calcium acetate (aq)

The quantity, quality and fat-content from the various milk samples differ with
the type of particular mammals and their fodder. The composition of milk
varies according to the animals from which it comes, providing the correct
growth rate and development for the young of that species. Casein is a slow

17 | Page
digesting protein and it is suspended in the milk in a complex called micelle. The
micelles are spherical and are 0.04 to 0.03 m in diameter. Milk composition
varies with the stage of location, age and breed. Milk is colloidal in nature due to
the presence of proteins. The proteins are heavy molecules; they form colloids
when dispersed in water medium. The primary function of protein in living cells
is to promote growth and maintenance. The nitrogen content of milk is
distributed among casein 76%, when protein and no nprotein nitrogen is 6%.The
structure of protein consists of a polypeptide chain of amino acids joined
together by peptide linkages.

Procedure:

 Take a clean dry beaker, and add 100 ml of cow’s milk to it. Heat the
apparatus till it reaches 50°C.
 To the warm milk add 10% acetic acid solution drop by drop with
constant stirring simultaneously. Fat along with casein will precipitate
out.
 Filter the solution and transfer the precipitate into a watch glass.
 To the precipitate, 40 ml of ethanol is added and mixed well. This
helps in dissolving fat in the ethanol solution leaving out only casein.
 Filter the
Weigh thesolution
dry solidagain
massand allow the precipitate to dry.


Repeat the experiment with other samples of milk

18 | Page
Observation:

S.no Samples Weight of milk Weight of casein Percentage of

used [100ml] extracted or precipitated casein

1 115.45g 8.260g 7.15%

Buffalo
milk

2 Cow milk 178.3g 8.860g 4.96%

3 Goat milk 115.45g 8.050g 6.97%

The various milk samples of goat milk, cow milk and buffalo milk contain 6.97% ,
4.96% and 7.15% of casein respectively. This shows that buffalo milk has the
highest content of casein protein present in it than cow milk or goat milk. The
lower amount of casein in cow milk may be due to the dominance of other
prevalent proteins and goat milk has minimal amount of goat milk but it has
almost an equivalent percentage of casein to that of cow milk.

Therefore, the order follows: Buffalo milk > Goat milk > Cow milk

COW MILK’S EXTRACTED CASEIN

19 | Page
BUFFALO MILK’S EXTRACTED CASEIN

GOAT MILK’S EXTRACTED CASEIN

20 | Page
 CONCLUSION
Through this project, my teammates and I have been able to determine the
amount of casein in different samples of milk. The amount of casein in cow
milk, buffalo milk and goat milk are as follows:
Buffalo milk > Goat milk > Cow milk
According to our hypothesis we assumed that cow milk will have the highest
casein content due to its daily consumption. However, experimental results
proved that our assumption is incorrect and buffalo milk has the highest casein
content. Despite having the highest casein content, buffalo milk is not
generally preferred owing to its high fat content. As fat is not easily digested,
people prefer other alternatives for daily consumption.
Studies suggest that goat milk can lead to weight gain due to saturated fats
present, so it needs to be consumed carefully. Goat milk has a certain lactose
which is hard to digest by humans and results in cramps, bloating, vomiting,
etc. Also, from a practical perspective, goat milk does not separate and the
cream knits with the milk.
Hence, cow milk is generally preferred by many households.

21 | Page
 APPLICATIONS
Casein protein in milk has various applications across several industries. The
major uses include food preparation, the medical industry, cosmetics, and
dietary supplements. The minor applications of this protein include shoe
polishes and cleaners, dressing of leather, printing, and sizing of textile,
soapmaking, and several others. In many cases, casein also serves as a binder,
emulsifying agent, or colloid. The major uses of casein proteins are as follows:

Paper Coatings:

The art papers and books are coated with pigmented casein. This provides a
suitable surface to create the half-tone illustrations. For this, acid casein having
an alkaline solution is mixed with the water slurry. To increase the water
resistance, lime or formaldehyde is also added. Further, the spread of the
mixture takes place through a coating machine, and it is dried.

Glues: Casein glue is used to form wood joints. These withstand the dampness
just for some duration. However, the prepared casein glue is in a powdered
form. Its composition takes place through lime, suitable sodium salts, acid
casein, and lastly, a fungicide. Before using this glue, this powder is dissolved in
water. Not just this, there are several other uses which casein proteins serve.
These mainly include paints, plastics, and human-made fibres.

22 | Page
 FUTURE PROSPECTS
Casein can be used as a potential carrier for biologically active agents and can be
utilized as encapsulation material. Caseins, being dietary proteins, are
considered to be GRAS [Generally recognised as Safe] and are expected to be
biocompatible, biodegradable and bioresorbable on oral administration. Their
easy access and relatively high concentration in the raw material from which
they are obtained, i.e. milk, make caseins inexpensive. Production of caseins is
well known, long established and considered simple. Their flexible and unfolded
conformation provides them with resistance to denaturation, which is beneficial
during processing and manufacture. The behaviour of casein micelles is driven by
changes of pH, causing them to shrink in an acidic environment. This
phenomenon is anticipated to confer protection of substances encapsulated in
casein during passage through the harsh environment of the stomach. The open
structure of caseins makes them vulnerable to proteolytic digestion, which in
some cases can be beneficial, such as for complete release of encapsulated
substances, but disadvantageous in others, e.g. unchecked burst release, but can
be controlled by utilizing crosslinking agents. This is probably due to rigidisation
of protein structure and blocking of residues recognised by proteolytic enzymes.
Caseins have also been shown to penetrate the plasma membrane in an energy-
independent fashion, ensuring enhanced cellular uptake of encapsulated
substances.
Casein has been used for both microencapsulation and nanoencapsulation of
biologically active agents, including pharmaceuticals, probiotic cells,
nutraceuticals and nutrients. Encapsulation approaches for pharmaceuticals
differ from methods used for other substances, but this does not limit their

23 | Page
potential for future work. Indeed, many approaches tested for nutraceuticals
were eventually used for pharmaceuticals.
The remarkable gelation properties of casein have been used to prepare
microcapsules that can encapsulate cells of probiotic bacteria. This process was
facilitated by the enzymatic action of rennet or transglutaminase. However,
additional work is necessary to improve the survival rate and stability of cells
during storage. Much progress has also been made on coacervation of casein
with polysaccharides to encapsulate hydrophobic substances. This process can
be achieved by adjusting the pH to low values, as caseins gain positive charge
in acidic pH, and can thus interact with the negative charge on polysaccharides.
However, such methods are probably limited to acid-insensitive substances, as
others may be damaged.
Caseins are naturally designed to bind and encapsulate high concentrations of
calcium phosphate, via self-assembly into micelles. This, combined with the
block distribution of hydrophobic regions of caseins, enables them to interact
with lipophilic substances, providing a basis for nanoencapsulation of
hydrophobic substances. In brief, particular factors are utilized to disrupt the
micellar structure, hydrophobic substances are added and bind to caseins,
followed by removal of disrupting factors and subsequent reassembly of
caseins into micelles with the substances of interest entrapped inside. Such
nanoparticles can provide solubilisation of lipophilic compounds in waterbased
solutions, in addition to improved bioavailability and protection against certain
conditions. Recently, self-assembly was also studied for isolated βcasein.

24 | Page
This biopolymer has a deblock structure in terms of charge and hydrophobicity
distribution along the chain, providing the foundation for its spontaneous
micellization. From 0 to 30 °C, β-casein changes from monomeric to micellized.
This temperature-driven process in such a mild thermal range suggests great
potential for nanoencapsulation of sensitive compounds.
Different casein nanoparticles have been shown to protect their contents
against cold (storage and lyophilisation), heat, oxidation, UV radiation, high
hydrostatic pressure processing and changes of pH and ionic strength.

Curcumin encapsulated with casein Nanoparticles

LIMITATIONS OF THE STUDY

• Casein from adulterated milk was not possible due to very less amount
present.
• The research was restricted to only three sources of milk to extract the
casein.

25 | Page
 BIBLIOGRAPHY

 https://www.verywellhealth.com/casein-5081318

 https://healthyeating.sfgate.com/functions-protein-
casein-10637.html

 https://www.healthline.com/nutrition/what-is-
casein#what-it-is
 https://www.britannica.com/science/casein
https://www.sciencedirect.com/topics/agricultur al-
and-biological-sciences/casein

 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5511
616/

26 | Page